HIV efficiently develops resistance to antiviral agents over time. Thus, there is an urgent need for better agents to treat HIV, especially drug-resistant strains of HIV. Recently, D-FDOC, (-)-beta-D-5-fluoro-1-[2-(hydroxymethyl)-1,3-dioxolan-4-yl]cytosine, was shown to have potent in vitro activity against drug-resistant strains of HIV (EC50 against HIV-1LAI is 0.04, mu M) without producing cross-resistance with 3TC, AZT or nevirapine. D-FDOC is also effective against HBV in culture. In primary mouse bone marrow cells, D-FDOC demonstrated no increase in lactic acid production even at 300 mu M. In contrast, treatment with L-FDOC resulted in a > 300-fold increase relative to untreated control. It is important to prepare enantiomerically pure D-FDOC to avoid potential toxicity from contaminated L-FDOC in the synthesis. However, pre-clinical and clinical development of D-FDOC has been hampered by the lack of an efficient procedure for large-scale preparation of this nucleoside. Thus, during Phase I, we will develop facile methods for the large-scale synthesis of D-dioxolane nucleosides. This chemistry will be useful not only to Pharmasset, but to other companies developing dioxolane nucleosides such as (-)-beta-D-2,6- diamino-9-[2-(hydroxymethyl)-1,3-dioxolan-4-yl]purine (DAPD) by Triangle Pharmaceuticals. Ohrui et al reported that 2'-deoxy-4'-C-ethynyl-ribo-nucleosides showed potent activity against HIV-1 in vitro with EC50 values ranging from 0.0003-0.03 mu M. 2'-C-Methyl-ribo-pentofuranosylcytosine was also shown to have potent anti-flavivirus activity. Based on these findings, we propose to design and synthesize novel racemic dioxolane nucleosides, in particular 2'-C-ethynyl and 5'-C-methyl substituted dioxolane nucleosides as antiviral agents for hepatitis C virus (HCV) and/or HIV in Phase I. Based on results from the biological testing of these compounds, we will then identify more active enantiomers for synthesis using the new process chemistry. In Phase II, we will apply our best methodology for large-scale synthesis of D-FDOC as well as the most promising new dioxolane nucleoside for further development towards an IND application.